Secretion of prostaglandin F2α at the time of maternal recognition of pregnancy leads to embryonic loss. Early embryonic losses are one of the important causes of repeat breeding which increases the calving interval and leads to economic losses to farmers. Non-steroidal anti-inflammatory drugs inhibit the synthesis of PGF2α. The present study was designed to study the effect of treatment by GnRH analogue - Buserelin acetate, Progesterone (P4) and Tolfenamic acid on 32 repeat-breeding crossbred cattle aged 3-8 years.
Int.J.Curr.Microbiol.App.Sci (2020) 9(5): 2630-2637 International Journal of Current Microbiology and Applied Sciences ISSN: 2319-7706 Volume Number (2020) Journal homepage: http://www.ijcmas.com Original Research Article https://doi.org/10.20546/ijcmas.2020.905.301 Evaluation of the Effect of GnRH Analogue, Progesterone and Tolfenamic Acid on Serum Progesterone Profile and Conception Rate in Repeat Breeding Crossbred Cattle Sudhanshu Pratap Singh1*, Ankesh Kumar2, Prakrutik Prafulchandra Bhavsar3, Mukesh Sahu4, Praveen Kumar1 and Sushil Kumar5 Department of Veterinary Gynaecology and Obstetrics, 2Department of Veterinary Clinical Complex, 5Department of Veterinary Medicine, Bihar Animal Sciences University, Patna, Bihar, India College of Veterinary Science and Animal Husbandry, Anand Agricultural University, Anand, Gujarat, India Department of Veterinary Gynaecology and Obstetrics, College of Veterinary Animal Sciences, G B Pant University of Agriculture and Technology, Pant Nagar, Uttarakhand, India *Corresponding author ABSTRACT Keywords Repeat breeders, Progesterone, Tolfenamic acid, Conception rate Article Info Accepted: 23 April 2020 Available Online: 10 May 2020 Secretion of prostaglandin F2α at the time of maternal recognition of pregnancy leads to embryonic loss Early embryonic losses are one of the important causes of repeat breeding which increases the calving interval and leads to economic losses to farmers Non-steroidal anti-inflammatory drugs inhibit the synthesis of PGF2α The present study was designed to study the effect of treatment by GnRH analogue Buserelin acetate, Progesterone (P4) and Tolfenamic acid on 32 repeat-breeding crossbred cattle aged 3-8 years All the animals were randomly divided into equal groups (n=8) Group was taken as positive control i.e inseminated on spontaneous estrous without any therapy Group animals were treated with injection Buserelin acetate @ 20 µg intramuscular (IM) at the time of artificial insemination (AI) Group animals were treated with inj Buserelin acetate @ 20 µg intramuscular at the time of AI followed by Injection P4 @100mg IM on days 4, 5, after AI Group animals were treated with inj Buserelin acetate @ 20 µg IM at the time of AI, inj P4 @ 100mg IM on days 4, 5, and inj Tolfenamic acid @4 mg/kg body weight IM on days 16, 17, 18 after AI Blood samples were collected from all the cows at day 0, 7, 16, 17, 18 of the estrous cycle for the study of serum progesterone profile The conception rate observed in groups 1, 2, and were 12.5%, 37.5%, 50% and 75% respectively Highest conception was observed in group in which, combinations of Buserelin acetate, Exogenous P and Tolfenamic acid was administered The study concluded that combination of Buserelin acetate, Exogenous P and Tolfenamic acid therapy helps to maintain the P4 level and significantly increases the conception rate by times than control group Introduction The bovine conceptus produces interferon-tau (IFNτ) which prevents luteolysis In addition to that functional corpus luteum (CL) produces more progesterone which is essential to supports pregnancy (Spencer and Bazer, 2002) There are two crucial periods of bovine pregnancy The first period is the first week of after breeding and second is day to 2630 Int.J.Curr.Microbiol.App.Sci (2020) 9(5): 2630-2637 28, when maternal recognition of pregnancy (MRP) takes place Approximately 32% of total embryonic loss occurs in this period (Wiltbank et al., 2016) This leads to a repeat of oestrous Repeat breeding (RB) is a considerable problem in cattle breeding which leads to large economic losses due to more inseminations, increased calving interval and increased culling rates Repeat breeding has been defined as the failure to conceive from or more regularly spaced services in the absence of detectable abnormalities (Bartlett et al., 1986) Several factors like nutritional stress (Bender et al., 2014), heat stress (Sakatani, 2017), transportation stress (Merrill et al., 2007) or other stress promotes secretion of prostaglandin F2α from the uterine endometrium This can cause lysis of the functional corpus luteum and leads to early embryonic death (Hockett et al., 2004) The two cyclooxygenase (COX) enzymes convert arachidonic acid into prostaglandin-H2 which is further converted into PGF2α through the enzyme prostaglandin-F-synthase The NSAIDs exhibits anti-inflammatory activity mainly based on the inhibition of the cyclooxygenase (COX) enzyme, results in inhibition of prostaglandin synthesis (Malm and Borisch, 2015) Several studies evaluated the effects of NSAIDs like Flunixine meglumine (Kasimanickam et al., 2019, 2018) and meloxicam (Amiridis et al., 2009; McDougall et al., 2016) Administration of the NSAID tolfenamic acid has significantly improved embryo transfer rate and pup delivery in mice (Schlapp et al., 2015) Administration of GnRH or GnRH analogue before artificial insemination induce preovulatory LH (luteinizing hormone) surge which controls ovulation, or post inseminations with supplementation of exogenous P4 to support early embryonic development (Amiridis et al., 2009) At days to post-ovulation, the embryos usually enter the uterus, undergoing genomic activation and increases in P4 concentration; therefore, this may be a physiologically important time in the cattle, so the administration of a low dose of P4 on days 4, and of the oestrous cycle increase the conception rate among repeat breeder cattle (Ferguson et al., 2012) This extends the life span of the bovine corpus luteum (CL) so this is one of the strategies aimed at reducing embryo loss by inhibiting the PGF2α in the endometrium during the critical period (Binelli et al., 2001; Pugliesi et al., 2011), Inhibition of PGF2α enhances the CL lifespan and avoiding detrimental and toxic effects of PGF2α on the embryo (Binelli et al., 2001) Materials and Methods Preparation of animals commencement of treatment before The study was conducted on 32 apparently healthy, 3-8 years old repeat breeding crossbred cattle All animals were dewormed with Fenbendazole @ 7.5 mg per kg body weight 60 days prior to the commencement of experiment and they were supplemented with 40 gm mineral mixture daily prior to experiment Grouping of animals and treatment Selected animals were randomly divided into four groups (each group containing animals) In group 1, animals were inseminated on spontaneous estrous without any treatment, in group 2, animals were treated with injection Buserelin acetate @ 20 µg IM at the time of artificial insemination (AI), in group 3, animal were treated with 2631 Int.J.Curr.Microbiol.App.Sci (2020) 9(5): 2630-2637 injection Buserelin acetate @ 20 µg IM at the time of AI followed by Injection P4 @100mg IM on days 4,5,6 after AI and in group 4, animals were treated with injection Buserelin acetate @ 20 µg IM at the time of AI, Injection P4 @100mg IM on days 4,5,6 and Inj Tolfenamic acid @4 mg/kg body weight IM On days 16,17,18 after AI All the animals were inseminated on spontaneous heat (Table 1) Statistical analysis of the effect of different treatments between the groups and within the group on different days and serum P4 concentration in of repeat breeder cows was studied by repeated measures ANOVA The multiple comparisons between group, day and interaction for different parameters were done by using Tukey test at 5% level of significance The analysis was done using JMP 9.0 software Blood collection and hormonal assay Results and Discussion Assuming estrous day as day 0, blood samples was collected in clot activator from all the cows at day 0, 7,16,17,18 of the cycle to harvest for hormonal estimation (Table 1) Level of P4 in blood was estimated by RadioImmuno Assay (RIA) Progesterone essay kit (M/S Beckman Coulter, Brea, CA) catalogue no IM 1188 as per the suggested protocol In brief all reagents were brought to the room temperature (20-25°C) before assaying Conception rate For estimation of serum progesterone, 50µl standard and 50µl serum sample were taken into antibody coated tubes 500µl 125Ilabelled serum progesterone was added to each antibody-coated tube Two additional non-coated tubes were prepared for total activity computation containing 500µl tracer (125I-labelled serum progesterone) and was set-aside until counting The contents of the tubes were mixed with a vortex and incubated for hr for serum progesterone estimation at room temperature, while continuously shaking (300-350 rpm) The incubation mixture was carefully aspirated while aspirator tip touched the bottom of the antibody coated tube so that all the liquid was removed The radioactivity was measured with SR-300 fully automatic gamma counter The conception rates observed in the present work in all four groups were 12.5%, 37.5%, 50% and 75% respectively in group 1, group 2, group and group The highest conception rate was observed in group In a similar study by Amiridis et al., (2009) reported highest conception rate of 33.76 % in repeat breeder cows which is lower conception rate than the present study (Aguiar et al., 2013) they reported conception rate of 66.70% with the use of meloxicam- a NASID and this is comparable with the present finding while they obtained 49% Conception rate in the control group which is higher than the present report Archbald et al., (1993) used GnRH at the time artificial insemination and reported the conception rate ranging from 33% - 40% this is comparable to the present finding (Fig 2) While using progesterone in a low dosage by (Ferguson et al., 2012) in an experiment observed conception rate of 45% which slight lower than the present finding Highest conception rate in present study was observed in group i.e 75%, where treatment had been given with combinations of GnRH analogue (Busereline acetate), Exogenous progesterone and non-steroidal anti- 2632 Int.J.Curr.Microbiol.App.Sci (2020) 9(5): 2630-2637 inflammatory drug (Tolfenamic acid) All the treatments were given at different days of estrus cycle when the chances of conception failure are more i.e Buserelin acetate on day when time of ovulation depend on concentration of LH which is govern by release of GnRH, then Progesterone injected on day 4, and when embryo travels from horn to uterus and Tolfenamic acid was given on day 16, 17 and 18 when MRP process are going on Least conception rate observed in group in which no treatment was given and kept as a control The combined administration of GnRH, Progesterone, and NSAID was effective in treatment of the repeat breeder cows Serum progesterone profile In group The mean total P4 concentration on day 0, 7, 16, 17 and 18 was 0.31±0.06, 2.31±0.27, 3.99±0.23, 4.63±0.29 and 4.40±0.31 ng/ml, respectively The significant (p